This is the fourth year of operation of the Section on Neuronal Circuitry. The focus this year has been generating data for scientific presentations and publications. Projects fall into two major categories: 1) Intrinsic electrical properties and synaptic inputs of medical olivocochlear (MOC) neurons in the brainstem, and 2) Influence of synaptic outputs of MOC neurons in the cochlea. Synaptic inputs of olivocochlear neurons in the brainstem and synaptic outputs in the cochlea Medial olivocochlear (MOC) neurons have cell bodies in the brainstem, where they receive excitatory synaptic inputs conveying sound information from the cochlea via the cochlear nucleus. Very little is known about the neuronal circuitry driving activity of MOC neurons because their cell bodies are difficult to locate in un-stained brain preparations. An initial goal of the lab is to develop and characterize a reliable genetic technique to perform single cell patch-clamp recordings from the MOC neurons in brain slices from mice. Using this technique, we aim to understand the diversity of synaptic inputs of MOC neurons to fully understand how the neurons are activated and modulated during auditory perception. We also aim to determine how the MOC neuron baseline electrical properties may change to contribute to neuronal hyperactivity that has been shown to occur in pathological situations such as in tinnitus or hyperacusis. Experimental accomplishments include development of a computational model of MOC neurons, including addition of accurate post-synaptic current waveforms and intrinsic electrical conductances from experimental data collected in our own laboratory. We have demonstrated the presence of the hyperpolarization-activated current Ih in MOC neurons and have incorporated it into the computational model to investigate the role of Ih in integration of synaptic inputs in the MOC neuron. In a second project we have discovered a pathway of sound-driven inhibition of MOC neurons and are characterizing the effect of this inhibition on MOC neuron activity. These experiments will elucidate the mechanisms of synaptic activation and inhibition of MOC neurons, which in turn drives their inhibition of mechanical activity in the cochlea, thus shaping the cochlear response to sound. A publication to describe our initial findings is in revision. Synaptic outputs of olivocochlear neurons in the cochlea MOC neurons project to the cochlea, where they decrease the mechanical movement of the basilar membrane by inhibiting cochlear OHCs. OHC activity enhances signaling to cochlear IHCs and shapes cochlear tuning curves and gain. MOC synapses onto OHCs are implicated in inhibiting OHC via coupling of cholinergic channel coupling to an SK potassium conductance. This effect is implicated in improved hearing in background noise, and protection of the cochlea against sound trauma. MOC neurons are also thought to release other neurotransmitters in the cochlea with poorly described effects on cochlear function. Equipment for this project has been installed and tested, and preliminary data is being collected. A post-doctoral fellow has recently been recruited and is currently in training to conduct experiments including patch-clamp electrophysiology and imaging in dissected cochlear preparations. VGLUTs in OHCs, and the central innervation patterns of type II cochlear afferent neurons In a collaborative project with Dr. Rebecca Seal at the University of Pittsburgh, the specific vesicular glutamate transporters (VGLUTs) employed by OHCs to load glutamate into presynaptic vesicles for release onto spiral ganglion afferents was determined. This led to generation of mutant mice lacking the VGLUTs from either inner hair cells (IHC) or OHC, or both. Use of these mutant mice allows isolation of afferent signaling by either type of hair cell, in order to determine the unique contribution that each pathway makes to perception of sound. We used these different mouse lines in an experiment in which the animals were exposed to noise, then their cochlear nuclei assessed for activation of the immediate early gene c-Fos. We determined that the OHC-type II afferent pathway can indeed respond to acoustic signals at non-pathological levels of sound, and evoke activation of neurons in the brainstem. A paper describing this work is currently in preparation.